Biswas Sondip K, Jiang Jean X, Lo Woo-Kuen
Department of Neurobiology, Morehouse School of Medicine, Atlanta, GA 30310, USA.
Mol Vis. 2009 Aug 4;15:1492-508.
To investigate the structural remodeling in gap junctions associated with cholesterol redistribution during fiber cell maturation in the adult chicken lens. We also evaluated the hypothesis that the cleavage of the COOH-terminus of chick Cx50 (formerly Cx45.6) during fiber cell maturation would affect the gap junction remodeling.
Freeze-fracture TEM and filipin cytochemistry were applied to visualize structural remodeling of gap junction connexons associated with cholesterol redistribution during fiber cell maturation in adult leghorn chickens (42-62 weeks old). Freeze-fracture immunogold labeling (FRIL) was used to label the specific Cx50 COOH-terminus antibody in various structural configurations of gap junctions.
Cortical fiber cells of the adult lenses contained three subtypes of cholesterol-containing gap junctions (i.e., cholesterol-rich, cholesterol-intermediate, and cholesterol-poor or -free) in both outer and inner cortical zones. Quantitative studies showed that approximately 81% of gap junctions in the outer cortex were cholesterol-rich gap junctions whereas approximately 78% of gap junctions in the inner cortex were cholesterol-free ones. Interestingly, all cholesterol-rich gap junctions in the outer cortex displayed loosely-packed connexons whereas cholesterol-free gap junctions in the deep zone exhibited tightly, hexagonal crystalline-arranged connexons. Also, while the percentage of membrane area specialized as gap junctions in the outer cortex was measured approximately 5 times higher than that of the inner cortex, the connexon density of the crystalline-packed gap junctions in the inner cortex was about 2 times higher than that of the loosely-packed ones in the outer cortex. Furthermore, FRIL demonstrated that while the Cx50 COOH-terminus antibody was labeled in all loosely-packed gap junctions examined in the outer cortex, little to no immunogold labeling was seen in the crystalline-packed connexons in the inner cortex.
Gap junctions undergo significant structural remodeling during fiber cell maturation in the adult chicken lens. The cholesterol-rich gap junctions with loosely-packed connexons in the young outer cortical fibers are transformed into cholesterol-free ones with crystalline-packed connexons in the mature inner fibers. In addition, the loss of the COOH-terminus of Cx50 seems to contribute equally to the transformation of the loosely-packed connexons to the crystalline-packed connexons during fiber cell maturation. This transformation causes a significant increase in the connexon density in crystalline gap junctions. As a result, it compensates considerably for the large decrease in the percentage of membrane area specialized as gap junctions in the mature inner fibers in the adult chicken lens.
研究成年鸡晶状体纤维细胞成熟过程中与胆固醇重新分布相关的缝隙连接结构重塑。我们还评估了一种假说，即纤维细胞成熟过程中鸡Cx50（原Cx45.6）羧基末端的裂解会影响缝隙连接重塑。
应用冷冻蚀刻透射电子显微镜和菲律宾菌素细胞化学方法，观察成年来亨鸡（42 - 62周龄）纤维细胞成熟过程中与胆固醇重新分布相关的缝隙连接连接子的结构重塑。冷冻蚀刻免疫金标记（FRIL）用于在缝隙连接的各种结构构型中标记特异性Cx50羧基末端抗体。
成年晶状体的皮质纤维细胞在皮质外层和内层区域均含有三种含胆固醇的缝隙连接亚型（即富含胆固醇、胆固醇含量中等以及胆固醇含量低或不含胆固醇）。定量研究表明，外层皮质中约81%的缝隙连接为富含胆固醇的缝隙连接，而内层皮质中约78%的缝隙连接为不含胆固醇的缝隙连接。有趣的是，外层皮质中所有富含胆固醇的缝隙连接显示连接子排列松散，而深层区域不含胆固醇的缝隙连接则呈现紧密的六边形晶体排列的连接子。此外，虽然外层皮质中作为缝隙连接特化的膜面积百分比约为内层皮质的5倍，但内层皮质中晶体排列的缝隙连接的连接子密度约为外层皮质中排列松散的连接子的2倍。此外，FRIL显示，虽然在外层皮质中检查的所有排列松散的缝隙连接中均标记了Cx50羧基末端抗体，但在内层皮质中晶体排列的连接子中几乎未见免疫金标记。
成年鸡晶状体纤维细胞成熟过程中缝隙连接经历显著的结构重塑。年轻的外层皮质纤维中连接子排列松散的富含胆固醇的缝隙连接在成熟的内层纤维中转变为连接子晶体排列的不含胆固醇的缝隙连接。此外，Cx50羧基末端的缺失似乎在纤维细胞成熟过程中对连接子从排列松散转变为晶体排列起到了同等作用。这种转变导致晶体缝隙连接中连接子密度显著增加。结果，它在很大程度上补偿了成年鸡晶状体成熟内层纤维中作为缝隙连接特化的膜面积百分比的大幅下降。
